PROJECT SUMMARY Despite continual improvements in surgical techniques, cancer recurrence after surgical resection remains a significant challenge in cancer therapy. It has also been verified that surgery can induce promotion of cancer metastasis. In this proposal, utilizing platelet as a delivery vehicle, the team intends to develop a transformative platform for locally releasing immune checkpoint inhibitors toward post-surgical eradication of residual cancer cells. In a preliminary study, using the B16F10 melanoma tumor-bearing C57BL/6 mouse model, it has been demonstrated that the anti-PDL1 (aPDL1) attached platelets (designated P-aPDL1) could facilitate the accumulation of aPDL1 toward the surgical bed where the residual microtumors remain. Importantly, the loaded aPDL1 can be effectively released from the activated platelets mediated by the platelet-derived microparticles (PMPs) upon in situ platelet activation. Moreover, platelets can generate a local inflamed tumor microenvironment, which could boost T cells activity as well as other immune cells. Here, the team proposes to further substantiate, optimize and extend the capability of platelets as a delivery platform for checkpoints blockade-based cancer immunotherapy. The team will validate the detailed treatment mechanism of P-aPDL1 as well as optimize its physicochemical property. The capability of P-aPDL1 for treating circulating tumor cells (CTCs) will also be evaluated. In addition, the team will evaluate the potential of platelets to achieve combination delivery of aPDL1 and gemcitabine (GEM), which can upregulate both PDL1 and PD1 on tumor cells and tumor infiltrating immune cells, respectively. Furthermore, the twam will extend this platform to co-deliver different ?cells?- therapeutics-loaded platelets and specific chimeric antigen receptor (CAR) T cells. Three aims will be pursued: in Aim 1, the capability of platelets for delivering checkpoint inhibitors will be validated and optimized; in Aim 2, the effectiveness of combination delivery of aPDL1 and GEM using platelets will be evaluated; in Aim 3, the innovative combination cell immunotherapy with platelets and CAR-T cells will be developed and assessed. The synergistic immune responses as well as systemic toxicity of this combination cells-based immunotherapy will be evaluated. The proposed research, when successfully demonstrated in human studies, would significantly enhance the anticancer efficacy and improving the patients? survival. This novel in situ bio-responsive strategy may also inspire new treatments applying bio-particulates for targeting and bio-responsive release of therapeutics.